Embolism formation during freezing in the wood of Picea abies

Freeze-thaw events can cause embolism in plant xylem. According to classical theory, gas bubbles are formed during freezing and expand during thawing. Conifers have proved to be very resistant to freeze-thaw induced embolism, because bubbles in tracheids are small and redissolve during thawing. In contrast, increasing embolism rates upon consecutive freeze-thaw events were observed that cannot be explained by the classical mechanism. In this study, embolism formation during freeze-thaw events was analyzed via ultrasonic and Cryo-scanning electron microscope techniques. Twigs of Picea abies L. Karst. were subjected to up to 120 freeze-thaw cycles during which ultrasonic acoustic emissions, xylem temperature, and diameter variations were registered. In addition, the extent and cross-sectional pattern of embolism were analyzed with staining experiments and Cryo-scanning electron microscope observations. Embolism increased with the number of freeze-thaw events in twigs previously dehydrated to a water potential of -2.8 MPa. In these twigs, acoustic emissions were registered, while saturated twigs showed low, and totally dehydrated twigs showed no, acoustic activity. Acoustic emissions were detected only during the freezing process. This means that embolism was formed during freezing, which is in contradiction to the classical theory of freeze-thaw induced embolism. The clustered pattern of embolized tracheids in cross sections indicates that air spread from a dysfunctional tracheid to adjacent functional ones. We hypothesize that the low water potential of the growing ice front led to a decrease of the potential in nearby tracheids. This may result in freezing-induced air seeding.     

Assessment of transpiration estimates for Picea abies trees during a growing season

Summary The tree-trunk heat balance method with internal heating and sensing of temperature was used to estimate sap-flow rate of spruce trees in a stand in southern Sweden. Sap-flow rate values were scaled up to stand transpiration and utilised for calculation of canopy conductance. The calculated values provided the basis for a function relating canopy conductance to vapour pressure deficit, which was implemented in the Penman-Monteith equation. The stand was mostly growing in non-limiting soil water conditions (irrigation regime applied during dry periods). The whole-season transpiration was assessed by two different approaches and then compared: the sap-flow rate measurements were scaled to stand transpiration and the adapted Penman-Monteith estimate. They gave similar results: the transpiration totals differed by 3% and the coefficient of determination of the linear regression was r² = 0.89. Similarly good was the assessment for a set of rainy days. The Penman-Monteith estimate adapted in this way proved to be reasonably precise and reliable in this forest stand and usable even in wet conditions. The seasonal transpiration of the spruce stand was 392 mm according to the adapted Penman-Monteith equation. Mean daily transpiration was 1.8 mm and daily maximum transpiration was 4.8–4.9 mm as estimated by sap-flow rate measurements.     

Daylength and temperature during seed production interactively affect adaptive performance of Picea abies progenies

Adaptive traits in Picea abies (Norway spruce) progenies are influenced by the maternal temperatures during seed production. Here, we have extended these studies by testing the effects of maternal photoperiod and temperature on phenology and frost hardiness on progenies. Using eight phytotron rooms, seeds from three unrelated crosses were made in an environmental 2 x 2 factorial combination of long and short days and high and low temperatures. The progenies were then forced to cease growth rapidly at the end of the first growing season. An interactive memory effect was expressed the second growth season. Progenies from high temperature and short days, and from low temperatures and long days, started growth later in spring, ceased shoot growth later in summer, grew taller and were less frost hardy in the autumn than their full siblings from low temperatures and short days, and from high temperatures and long days. Norway spruce has developed a memory mechanism, regulating adaptive plasticity by photoperiod and temperature, which could counteract harmful effects of a rapidly changing climate.     

Variation in total and specific RNA during inwintering of two contrasting populations of Picea abies

Seedlings of a southern (Romanian) and a northern (Swedish) population of Picea abies (L.) Karst. were cultivated in a phytotron under continuous lighl and 20°C for 10 weeks. Subsequently they were exposed to a regime of long nights (16 h). initially at 20°C but with stepwise lowering to 10°C over 12 weeks. Samples of needles were taken for total RNA extraction and studies of gene expression at intervals from just before the change of regime and onwards. At the same time samples were taken for abseisic acid (ABA) determination. Extension growth, degree of budrest and frosl tolerance were also recorded. The main results were as follows. (1) Extractable total RNA per g fresh weight of needles increased by 150–250% through the period of long nights. (2) The capacity of the total RNA to direct protein synthesis in a cell-free system declined in response to long nights to a minimum value of 5% (northern population) and 23% (southern population) of the reference values under continuous light. (3) In vitro translatability was highly correlated with the size of the polyadenylated fraction of the total RNA, which declined also when normalized to needle fresh weight rather than total RNA. (4) A peak of abscisic acid content in the southern population at 3 days into the long night treatment coincided with a trough in the in vitro translatability of the total RNA. (5) The steady state levels of rbc S-mRNA as a fraction of total RNA declined rapidly to a final value of about 15% of the reference value in both populations, the decline being more rapid for the northern population. (6) In contrast. ubiquitin-mRNA showed an initial increase, particularly in the northern population, and at the end of the long night treatment was only 40% under the reference value. (7) Rapid changes in the molecular characters occurred during the first 3–4 weeks when the morphological and physiological changes were most rapid.     

Abscisic acid content at defined levels of bud dormancy and frost tolerance in two contrasting populations of Picea abies grown in a phytotron

Seedlings of a southern (Romanian) and a northern (Swedish) population of Picea abies were cultivated under continuous light and 20°C for 10 weeks. To arrest growth, induce terminal bud dormancy and promote frost tolerance the seedlings were then exposed to 16 h nights for 12 weeks, with gradually lower temperature during the last 6 weeks. Samples for estimating the abscisic acid content of the needles were taken just before the onset of the night treatment, at day 3 of the treatment, and then with one, and later 2 week, intervals. From the second week onwards (third week for frost tolerance) bud dormancy and frost tolerance were assessed at the same time as abscisic acid (ABA) determinations. Phosphate-buffered saline extracts were purified on mini-columns (in some cases immunoaffinity colums) and quantified by HPLC. The degree of dormancy was estimated by transferring the seedlings to growth conditions and determining the number of days until growth was resumed. The frost tolerance of the needles exposed to –10°C and –20°C was classified in 6 classes. The frost tolerance of the terminal buds was estimated as the number of seedlings that showed some growth after 6 weeks in growth conditions. The night treatment rapidly induced terminal bud dormancy in both populations, but the release of dormancy occurred earlier in the northern population. The needles and the terminal buds became highly frost tolerant more rapidly in the northern than in the southern population and before the temperature decrease. The degree of dormancy began to decline before full frost tolerance was obtained in the southern population and this decline continued in both populations, while frost tolerance remained at a high level. The southern population showed a transient peak in ABA content at day 3. Although the ABA content of the northern population was lower than in the southern before the 16-h night treatment, it increased in the northern population during the treatment period, in particular after the temperature decrease.     

Diagnosis of damage to Norway spruce (Picea abies) through biochemical criteria

Bioindication can be carried out at different hierarchical levels, eg. cell, organism, and ecosystem. While the monitoring of damage by visible criteria (e.g. loss of needles) is connected with the organism as a whole, the monitoring of damage by biochemical indicators is above all connected with cell metabolism.
The degree of vitality of a tree can be ascertained through the integration of a number of biochemical parameters. Furthermore, a differential diagnosis of a particular stress pattern can be carried out because of the feedback pattern of several biochemical indicators. In order to describe and interpret biochemical or physiological changes that have been caused by a number of factors, multivariate statistical methods are being used more frequently. Apart from cluster and discriminant analysis, it is especially factor analysis which provides a helpful tool when dealing with problems in the field of environmental analysis. Factor analysis can be used for an integrating as well as a differentiating assessment.
Within the framework of forest damage research, numerous changes at the level of cell metabolism have been detected to which a bioindicative character can be attached. A number of physiological and biochemical parameters with bioindicative character concerning Norway spruce are presented.     

Chemical and cytological changes during the autolysis of yeasts

Summary Cell suspensions ofSacharomyces cerevisiae, Kloeckera apiculata andCandida stellata were autolyzed in phosphate buffer, pH 4.5, for up to 10 days. Cell dry weights decreased by 25–35% after 10 days. Based on initial cell dry weight, the soluble autolysate consisted of: carbohydrate (principally polysaccharide) 3–7%; organic acids 3–6%; protein 12–13%; free amino acids 8–12%; nucleic acid products 3–5%; and lipids 1–12%. The main organic acids in autolysates were propionic, succinic and acetic and the main amino acids were phenylalanine, glutamic acid, leucine, alanine and arginine. Approximately 85–90% of cellular RNA and 25–40% of cellular DNA were degraded during autolysis. Both neutral lipid and phospholipid components were degraded, with neutral lipids but not phospholipids being found in autolysates. Scanning and transmission electron micrographs showed retention of cell wall structure and shape during autolysis, but there was extensive intracellular disorganization withinS. cerevisiae andC. stellata. There were differences in the autolytic behavior ofK. apiculata compared withS. cerevisiae andC. stellata.This paper is dedicated to Professor Herman Jan Phaff in honor of his 50 years of active research which still continues.     

Shoot development of Norway spruce (Picea abies) involves changes in piperidine alkaloids and condensed tannins

Key Message

Secondary chemistry of P. abies peaks early in shoot development. Condensed tannins accumulate already in late buds while piperidine alkaloid biosynthesis take place in early stage shoots.

Abstract

Plants protect new vegetative parts with defensive secondary metabolite compounds. We investigated how concentrations of piperidine alkaloids and condensed tannins change during bud burst and shoot growth in adult Picea abies. We detected 12 individual piperidine compounds, of which epipinidinone and 1,2-dehydropinidinone and two tentatively identified 1,6-imines are reported for the first time in P. abies. In addition three piperidine alkaloid compounds remain partly identified. We found that concentrations of both total piperidine alkaloids and condensed tannins were highest immediately after bud burst. While concentrations of condensed tannins started to increase during bud opening, the dilution effect decreased concentrations in the developing needles of mature branches. By contrast, the decrease of total alkaloid concentrations in mature shoots was not due to the dilution effect, but was connected to the disappearance of precursor components of biosynthesis. The concentrations of major alkaloid components remain stable from dormant buds to mature needles and twigs, underlining their importance for P. abies, although their real ecological significance is yet to be solved. Based on the structural features and timing of appearance of individual compounds, we also propose a hypothetical biosynthesis route for trans-substituted coniferous piperidine alkaloids.     

Hormone levels in the cambial region of intact Picea abies during the onset of cambial activity

Microdialysis probes were used to sample the cambial region of Picea abies stems during the cambial reactivation period. The concentrations of ethylene and zeatinriboside in each sample were measured by gas chromatography and enzyme-linked immuno-sorbent assay, respectively. The ethylene level increased throughout the experimental period, except for a temporary decrease that occurred about the time of budburst on the lower branches. The zeatin-riboside level changed relatively little during the same period. The results indicate that increased activity in the cambial region was associated with an increase in the level of ethylene but not of zeatinriboside.     

Loss of HuR is linked to reduced expression of proliferative genes during replicative senescence

Cellular aging is accompanied by alterations in gene expression patterns. Here, using two models of replicative senescence, we describe the influence of the RNA-binding protein HuR in regulating the expression of several genes whose expression decreases during senescence. We demonstrate that HuR levels, HuR binding to target mRNAs encoding proliferative genes, and the half-lives of such mRNAs are lower in senescent cells. Importantly, overexpression of HuR in senescent cells restored a "younger" phenotype, while a reduction in HuR expression accentuated the senescent phenotype. Our studies highlight a critical role for HuR during the process of replicative senescence.     

Identification of a RAPD marker linked to the pendula gene in Norway spruce (Picea abies (L.) Karst. f. pendula)

The pendula phenotype of Norway spruce [Picea abies (L.) Karst f. pendula] is characterized by narrow crowns and strong apical dominance and is controlled by a single dominant gene (P). This defined genetic control presents one of the few opportunities to map a single gene controlling a morphological trait in a forest tree. We used random amplified polymorphic DNA (RAPD) markers and bulked segregant analysis to identify one locus OPH10_720, linked to the pendula gene. The estimated recombination frequency (r) between OPH10_720 and P was 0.046 (SE _r =0.032). Mapping of the pendula gene is an important first step towards the ultimate identification and cloning of this gene.     

Response of fine roots to an experimental gap in a boreal Picea abies forest

We examined the initial response of the quantity and distribution of fine roots to the creation of an experimental canopy gap with a diameter of 50 m in a mature managed Norway spruce forest. Under the canopy, the fine root length densities of trees, shrubs, and grasses and herbs were 3207, 707 and 2738 m m^–2, respectively. The fine root biomass of trees, shrubs, and grasses and herbs were 182, 47 and 52 g m^–2, respectively. Two growing seasons after gap creation hardly any fine tree roots were found in the middle part of the gap. The living tree roots in the gap edge zone were mainly located within a 5-m distance from the standing edge trees. The indices developed here to show the influence of trees on fine root lenght density clearly revealed the effect of the vicinity of living trees on fine root lenght density. The root densities of grasses, herbs and dwarf shrubs did not show a clear response to gap creation despite the increase of their foliage. Our results suggest that in boreal spruce forests a gap disturbance creates a distinct tree root gap and that the gap edge trees do not extend their root systems rapidly into the formed root gap.     

Characterization of embryogenic cell lines of Picea abies in relation to their competence for maturation

Summary Embryogenic cell lines of Picea abies are categorized into three groups (polar, solar, and undeveloped) based on the organization of the somatic embryos within the tissue and the ability of the somatic embryos to proceed through a maturation process when treated with ABA. The polar and the solar types consist of somatic embryos with densely packed embryonic regions subtended by vacuolated suspensors. Both types of tissue regenerate mature somatic embryos when treated with ABA. Almost all mature somatic embryos develop further into shoots or plantlets. The undeveloped type consists of somatic embryos comprised of only a few loosely aggregated cells in their embryonic regions. Mature somatic embryos were not observed with this tissue type.Abbreviations ABA cis-trans abscisic acid - A1 polar type - A2 solar type - B undeveloped type - BA benzyladenine - 2,4-D 2,4 di-chlorophenoxyacetic acid - LP von Arnolds medium (1987)